MicroSweat: A Wearable Microfluidic Patch for Noninvasive and Reliable Sweat Collection Enables Human Stress Monitoring

Abstract Stress affects cognition, behavior, and physiology, leading to lasting physical and mental illness. The ability to detect and measure stress, however, is poor. Increased circulating cortisol during stress is mirrored by cortisol release from sweat glands, providing an opportunity to use it as an external biomarker for monitoring internal emotional state. Despite the attempts at using wearable sensors for monitoring sweat cortisol, there is a lack of reliable wearable sweat collection devices that preserve the concentration and integrity of sweat biomolecules corresponding to stress levels. Here, a flexible, self‐powered, evaporation‐free, bubble‐free, surfactant‐free, and scalable capillary microfluidic device, MicroSweat, is fabricated to reliably collect human sweat from different body locations. Cortisol levels are detected corresponding to severe stress ranging from 25 to 125 ng mL−1 averaged across multiple body regions and 100–1000 ng mL−1 from the axilla. A positive nonlinear correlation exists between cortisol concentration and stress levels quantified using the perceived stress scale (PSS). Moreover, owing to the sweat variation in response to environmental effects and physiological differences, the longitudinal and personalized profile of sweat cortisol is acquired, for the first time, for various body locations. The obtained sweat cortisol data is crucial for analyzing human stress in personalized and clinical healthcare sectors.

S3 Figure S1. Demonstration of repeatability of laser cutting process. The repeatability of the optimized laser cutting parameters for fabrication of MicroSweat with different channel widths of a 250 µm (Scale bar: 10 mm), b 300 µm, c 400 µm, and d 450 µm.

SI.3. Evaluation of the Bracke model in the numerical modeling
To evaluate the Bracke model in this numerical modeling, the water flow in a straight microchannel with the width and height of 300 and 80 µm, respectively, was simulated and compared to the  Figure S5b shows this phenomenon for a long channel (20 mm in length) where the flow rate decreases from 200 mm/s to 30 mm/s. However, the channel length in MicroSweat is much smaller (3-6 mm in length) than that simple capillary channel, leading to a considerably lower decrease in the flow rate for only 30% of the initial flow rate, i.e., down to 120 mm/s. Although adding the sweat gland force to the capillary force further contributes to increasing the sweat flow rate, the capillary pressure alone is sufficient to fill up the MicroSweat patch.

SI.4. Sweat evaporation rate in Microsweat during human testing
The mass of water evaporated during human testing of MicroSweat was calculated for a minimum of 10 min exposure time for filling the last fiber and a maximum of 40 min exposure time for filling the first fiber. According to Langmuir's free evaporation equation in the vacuum, the mass loss rate at a given temperature T is obtained using the equation below [50,51] .
where dm/dt is the rate of mass loss (kg.s−1), S is evaporating surface area (πd2/4), d is the diameter of the orifice, which is the vent diameter in this case, Pv is vapor pressure (Pa), M is the molecular weight of the vapor of the evaporating compound (kg mol−1), R is the gas constant (JK −1 mol −1 ), T is the absolute temperature, and α is vaporization coefficient. In a vacuum, α is assumed to be 1, but as commonly found in TG experiments, α has a significantly different value in a flowing gas atmosphere. We consider α as 5.8-6.6 × 10 -5 (Pa -1 s -1 m 2 ) according to the studies with significant deviation from vacuum conditions. Therefore, the mass loss rate was estimated to be about 4.4 × 10 -10 g. s -1 , resulting in a mass loss value of 0.001 mg (0.001 µL) during a maximum of 40 min human tests. This amount can be compared to the amount of sweat absorbed by the for nitrocellulose fibers, showing that there is almost no sweat loss during the experiment. The assumptions here are: i) 99% of the sweat is water, 1 and ii) the rate of mass loss is calculated for the bulk of water. 2 Figure S7. Comparing the capacity of nitrocellulose fibers and glass microfibers' ability to absorb the sweat quantified using optical imaging.

SI.5. Saturation test of the fibers
A fluorescent dye dissolved in the water was injected into MicroSweat (using multiple pipettes delivering simultaneously to all inlets) until the fibers absorbed the dyed water ( Figure. S8a, b).
The transparency of this chip guarantees that all fluorescent light detected by the fluorescent microscope is stemmed from either the dyed water or the fiber without being affected by light reflection from PSA. The fibers also showed that they have no or minimal autofluorescent signal.

SI. 6. Order filling of MicroSweat simulated using a computational model
We further conducted a computational simulation to support the sequential filling of the storage chambers. Figure S9 shows the results of the ANSYS simulation, with two assumptions considered: a) the storage chambers are removed in this simulation, and b) only one channel collects the sweat at the inlet. Figure S9a shows the visual two-phase volume fraction indicating that the delay valves are activated sequentially from the bottom to the top. Figure S9b  S11 similar sequence of filling was also confirmed during human testing with the MicroSweat patch.
Given the assumptions implied, only the sequence but not the filling time values are comparable between the experimental data and numerical simulations in this simulation.

SI. 8. Perceived stress test
A more precise evaluation of personal stress can be determined by using a variety of instruments developed to measure the stress levels of individuals. A known method of measuring stress levels is the Perceived Stress Scale (PSS) as a classic stress assessment instrument. The PSS, initially developed in 1983, remains a popular choice for understanding different situations affecting human feelings and perceived stress. The PSS questions ask about the feelings and thoughts of the individual during the last month. The subject is asked to indicate how often he/she felt or thought a certain way. Individual scores on the PSS can range from 0 to 40, with higher scores indicating higher perceived stress. Scores ranging from 0-13 are considered low stress. Scores ranging from 14-26 are regarded as moderate stress. Scores ranging from 27-40 are considered high perceived stress. Figure S11. The standard curve of cortisol ELISA assay and the calibration curve used for detection of different concentrations of cortisol in spiked artificial sweat samples.